Mutations in germ cells are critical components of all hereditary diseases in humans. When present in somatic cells, mutations can constitute the essential pathogenetic basis of cancer, and of phenomena such as ageing. The potential for DNA damage to be translated into permanent mutations is mitigated by a complex repertoirs of cellular responses, including the process of nucleotide excision repair, where by damaged nucleotides are enzymatically excised from the genome of living cells. The long-term goals of this proposal are to understand the molecular mechanism of nucleotide excision repair of DNA in eukaryotic cells, using the yeast Saccharomyces cerevisiae as a model system. The proposed studies include the isolation of genes known from genetic studies to be required for nucleotide excision repair, by molecular cloning. The genes will be characterized in detail and tailored into suitable expression vectors for overexpression of proteins. These proteins will be purified from yeast cells with a view to reconstituting a damage-specific DNA incising activity in vitro and to characterizing the biochemistry of excision repair in yeast. The cloned yeast genes will also be expressed in mammalian cells known to be defective in nucleotide excision repair, with a view to demonstrating functional homology between DNA repair genes from lower and higher eukaryotes.